ROLE OF TISSUE MICRO ARCHITECTURE ON HEPATOCYTE FUNCTION

组织微结构对肝细胞功能的作用

• 批准号: 6050969
• 负责人: SANGEETA N. BHATIA
• 金额: $ 17.57万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-09-30 至 2004-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6050969
• 关键词: albumins; aminoacid analog; bioengineering /biomedical engineering; cell cell interaction; collagen; cytochrome P450; extracellular matrix; fibroblasts; gap junctions; laboratory rat; liver cells; liver function; membrane channels; mixed tissue /cell culture; proline; proteoglycan; tissue engineering; urea

Our overall goal is to develop highly functional hepatocellular tissue constructs that can be used in the treatment of liver failure. While there has been reasonable progress towards development of cell-based therapies for liver disease, the key determinants of high levels of liver- specific function in isolated hepatocytes have remained elusive. We and others have shown that co-cultivation of hepatocytes with non- parenchymal cells increases hepatocellular function in vitro; however, the fundamental modulators of hepatocellular function have yet to be clearly elucidated. The limitations in our understanding of relevant hepatocellular cues is, in part, due to the spatial complexity of multicellular constructs. We have shown previously that micropatterning (spatial localization of cells on solid substrates in patterns) can be utilized to control and study key cell-cell interactions. It is our hypothesis that the same cellular constituents in different spatial configurations will produce variations in the function of the resultant tissue. In this proposal, therefore, we aim to define the relationship between tissue architecture and tissue function in co-cultures of hepatocytes and non-parenchymal cells. A clear picture of the interplay between homotypic (hepatocyte/hepatocyte) interaction, heterotypic (hepatocyte/non- parenchymal cell) signaling, and cell-matrix interactions will be fundamental to the future design and implementation of hepatocyte-based tissue engineered medical products. Towards this end, our specific aims are: l). To determine the role of homotypic (hepatocyte/hepatocyte) interactions on hepatocellular function. Homotypic interactions will be varied (from single hepatocytes to multicellular islands), and cultures probed for markers of liver-specific function. The role of ECM cues and gap junctions- normally found at the homotypic interface- will also be examined in this context. 2). To investigate the role of heterotypic (hepatocyte/non-parenchymal cell) interaction on hepatocellular function. Co-cultures of rat hepatocytes and 3T3 fibroblasts will be micropatterned such that the spatial arrangement of both cell types is specified. We have previously shown that spatial configuration may influence hepatocyte function through either: amount of heterotypic interaction or shape of hepatocellular islands. These key parameters of tissue micro-architecture will be varied and co-cultures probed for markers of liver-specific function to isolate the determinants of tissue function. The role of ECM cues and heterotypic gap junctions-putative mediators of the co-culture 'effect' will be examined. 3).To optimize functionality of tissue constructs for tissue engineering applications. We will investigate the functional consequences of reduction in non-parenchymal number as well increased heterotypic interaction through use of 3-dimensional, grooved, micropatterned substrates. These studies will enable the formation of a fundamental framework that will guide the design and implementation of current and future hepatocyte-based medical therapies.

我们的总体目标是开发可用于治疗肝功能衰竭的高功能肝细胞组织结构。虽然以细胞为基础的肝病疗法的发展已经取得了合理的进展，但在分离的肝细胞中高水平的肝脏特异性功能的关键决定因素仍然难以捉摸。我们和其他人已经证明，肝细胞与非实质细胞共培养在体外提高了肝细胞的功能；然而，肝细胞功能的基本调节因素尚未被清楚地阐明。我们对相关肝细胞信号的理解的局限性，部分是由于多细胞结构的空间复杂性。我们之前已经证明，微图案化(细胞在固体基质上的空间定位)可以被用来控制和研究关键的细胞-细胞相互作用。我们的假设是，相同的细胞成分在不同的空间构型中会产生不同的组织功能。因此，在这项建议中，我们的目标是确定肝细胞和非实质细胞共培养中组织结构和组织功能之间的关系。清楚地了解同型(肝细胞/肝细胞)相互作用、异型(肝细胞/非实质细胞)信号和细胞-基质相互作用之间的相互作用，将是未来基于肝细胞的组织工程医疗产品设计和实施的基础。为此，我们的具体目标是：L)。目的：探讨同型(肝细胞/肝细胞)相互作用对肝细胞功能的影响。同型相互作用将是多种多样的(从单一肝细胞到多细胞岛)，培养将探索肝脏特异性功能的标记。ECM线索和缝隙连接的作用--通常在同型界面上发现--也将在这方面进行研究。2)。探讨异型(肝细胞/非实质细胞)相互作用对肝细胞功能的影响。大鼠肝细胞和3T3成纤维细胞的共培养将被微图案化，从而指定两种细胞类型的空间排列。我们先前已经证明，空间构型可能通过异型相互作用的数量或肝细胞岛的形状来影响肝细胞的功能。组织微结构的这些关键参数将是不同的，并将通过共培养探索肝脏特异性功能的标志，以分离组织功能的决定因素。ECM线索和异型缝隙连接的作用--假定的共培养‘效应’的中介--将被检验。3)为组织工程应用优化组织构建物的功能。我们将通过使用三维、凹槽、微图案化的衬底来研究减少非实质数量以及增加异型相互作用的功能后果。这些研究将形成一个基本框架，指导当前和未来基于肝细胞的医疗疗法的设计和实施。